The overall goal is to develop a treatment capable of correcting lysosomal storage diseases using the murine model of Mucopolysaccharidosis Type VII (MPS VII) which displays phenotypic characteristics similar to human patients.
The specific aims are to:
(Aim #1) Optimize retroviral transduction of hematopoietic stem cells by evaluating combinations of growth factors with and without a marrow stromal cell layer during pre- and cocultivation. This will be accomplished by incubating transduced marrow cells with a substrate for the introduced gene that produces a fluorescent product. The same cells are also fluorescently labeled with lineage specific antibodies. Image analysis is used to enumerate transduced non-lineage cells and determine efficiency. Further verification will be accomplished by in vivo engraftment and repopulation assays.
(Aim #2) Evaluate two new retroviral constructs containing beta-glucuronidase (beta-gus) cDNA by examining their ability to maintain long-term, high expression in vivo in adult hematopoietic stem cells and their progeny. The construct LCAGbetaGSN contains beta-gus driven by a beta-actin promoter/intron enhancer element. The second vector contains beta-gus driven by an LTR from the murine embryonic stem cell virus.
(Aim #3) Determine the therapeutic efficacy of in utero stem cell transplantation with retrovirally transduced hematopoietic cells. Information gathered from Aims #1 and #2 will allow optimal transduction and characterization of the most appropriate vector for in utero therapy. MPS VII recipients will be examined for amount and length of enzyme expression as well as correction of tissue pathology.
| Soper, B W; Lessard, M D; Vogler, C A et al. (2001) Nonablative neonatal marrow transplantation attenuates functional and physical defects of beta-glucuronidase deficiency. Blood 97:1498-504 |
| Soper, B W; Duffy, T M; Vogler, C A et al. (1999) A genetically myeloablated MPS VII model detects the expansion and curative properties of as few as 100 enriched murine stem cells. Exp Hematol 27:1691-704 |
| Soper, B W; Pung, A W; Vogler, C A et al. (1999) Enzyme replacement therapy improves reproductive performance in mucopolysaccharidosis type VII mice but does not prevent postnatal losses. Pediatr Res 45:180-6 |
